Porphyromonas gingivalis (formerly Bacteroides gingivalis) is an obligately anaerobic bacterium which is implicated in periodontal disease. P. gingivalis produces several distinct proteolytic enzymes, many of which are recognized as important virulence factors. A number of physiologically significant proteins, including collagen, fibronectin, immunoglobulins, complement factors C3, C4, C5, and B, lysozyme, iron-binding proteins, plasma proteinase inhibitors, fibrin and fibrinogen, and factors of the plasma coagulation cascade system, are hydrolyzed by P. gingivalis proteases. Broad proteolytic activity plays a role in the evasion of host defense mechanisms and the destruction of gingival connective tissue in progressive periodontitis.
Progressive periodontitis is characterized by acute tissue degradation promoted by collagen digestion and a vigorous inflammatory response characterized by excessive neutrophil infiltration. Gingival crevicular fluid accumulates in periodontitis as periodontal tissue erosion progresses at the foci of the infection, and numerous plasma proteins are exposed to proteinases expressed by the bacteria at the injury site. Neutrophils are recruited to the gingiva, in part, by the humoral chemotactic factor C5a. The complement components C3 and C5 are activated by complex plasma proteases with “trypsin-like” specificities called convertases. The human plasma convertases cleave the α-chains of C3 and C5 at a specific site generating biologically active factors known as anaphylatoxins (i.e. C3a and C5a). The anaphylatoxins are potent proinflammatory factors exhibiting chemotactic and/or spasmogenic activities as well as promoting increased vascular permeability. The larger products from C3 and C5 cleavage (i.e. C3b and C5b) participate in functions including complement cascade activation, opsonization, and lytic complex formation.
Recent studies have indicated that this periodontopathogen produces at least seven different enzymes belonging to the cysteine and serine catalytic classes of peptidases, among which three cysteine proteinases (gingipains) are predominant (Potempa, J., et al. (1995) Prospect. Drug Discovery and Design 2, 445-458). The gingipains are the best characterized group of P. gingivalis enzymes as their structure, function, enzymatic properties and pathological significance are known. From in vitro studies it is apparent that two gingipains R (also referred to generally as “Arg-gingipains” and more specifically as RgpA and RgpB), enzymes specific for cleavage at Arg-Xaa peptide bonds, have a significant potential to contribute to the development and/or maintenance of a pathological inflammatory state in infected periodontal pockets through: (i) activation of the kallikrein-kinin cascade, (ii) the release of neutrophil chemotactic activity from native and oxidized C5 of the complement pathway, and (iii) activation of factor X. In addition, gingipain K (also referred to as “Lys-gingipain”), an enzyme which cleaves Lys-Xaa peptide bonds, degrades fibrinogen. This may add to a bleeding on probing tendency associated with periodontitis. Finally, the presence of a hemagglutinin/adhesion domain in the non covalent multiprotein complexes of RgpA and gingipain K suggests participation of these enzymes in the binding of P. gingivalis to extracellular matrix proteins which may facilitate tissue invasion by this pathogen.
In comparison to the gingipains, relatively little is known about other cysteine proteinases produced by P. gingivalis. Two genes, referred to as tpr and prtT have been cloned and sequenced and although they encode a putative papain-like and streptopain-like cysteine proteinases, respectively, neither has been purified and characterized.
The presence of serine proteinase activity in cultures of P. gingivalis has been known for several years; however, only limited information is available about such enzymes. Indeed, a serine endopeptidase has been isolated from culture media, although it was only superficially characterized (Hinode D., et al., (1993) Infect. Immun. 59, 3060-3068). On the other hand, an enzyme referred to as glycylprolyl peptidase (DPP IV) was found to be associated with bacterial surfaces and two molecular mass forms of this peptidase have been described. This enzyme has also been shown to possess the ability to hydrolyze partially degraded type I collagen, releasing the Gly-Pro dipeptide, and it was suggested that, in collaboration with collagenase, glycylprolyl peptidase may contribute to the destruction of the periodontal ligament (Abiko, Y., et al. (1985) J. Dent. Res. 64, 106-111). In addition to this potential pathological function, glycylprolyl peptidase may also play a vital role in providing P. gingivalis with dipeptides which can be transported inside the cell and serve as a source of carbon, nitrogen, and energy for this asaccharolytic organism. Recently, a gene encoding glycylprolyl peptidase in P. gingivalis has been cloned and sequenced, and it is now apparent that this enzyme is homologous to dipeptidyl-peptidase IV (DPP-IV) from other organisms (Kiyama, M., et al. (1998) 1396, 39-46). The nucleotide sequence of the genome of this bacterium is currently being determined by The Institute for Genomic Research, and is available on the World Wide Web at tigr.org.